title: Measuring chromatin interaction dynamics on the second time scale at single-copy genes.
abstract: The chromatin immunoprecipitation (ChIP) assay is widely used to capture interactions between chromatin and regulatory proteins, but it is unknown how stable most native interactions are. Although live-cell imaging suggests short-lived interactions at tandem gene arrays, current methods cannot measure rapid binding dynamics at single-copy genes. We show, by using a modified ChIP assay with subsecond temporal resolution, that the time dependence of formaldehyde cross-linking can be used to extract in vivo on and off rates for site-specific chromatin interactions varying over a ~100-fold dynamic range. By using the method, we show that a regulatory process can shift weakly bound TATA-binding protein to stable promoter interactions, thereby facilitating transcription complex formation. This assay provides an approach for systematic, quantitative analyses of chromatin binding dynamics in vivo.
authors.affiliation: Department of Biochemistry and Molecular Genetics, University of Virginia Health System, Charlottesville, VA 22908, USA.
authors.forename: Kunal
authors.initials: K
authors.lastname: Poorey
authors.forename: Ramya
authors.initials: R
authors.lastname: Viswanathan
authors.forename: Melissa N
authors.initials: MN
authors.lastname: Carver
authors.forename: Tatiana S
authors.initials: TS
authors.lastname: Karpova
authors.forename: Shana M
authors.initials: SM
authors.lastname: Cirimotich
authors.forename: James G
authors.initials: JG
authors.lastname: McNally
authors.forename: Stefan
authors.initials: S
authors.lastname: Bekiranov
authors.forename: David T
authors.initials: DT
authors.lastname: Auble
mesh_terms: D000251:Adenosine Triphosphatases / Q000737:chemistry / Q000378:metabolism; D002843:Chromatin / Q000737:chemistry / Q000378:metabolism*; D047369:Chromatin Immunoprecipitation / Q000379:methods*; D003432:Cross-Linking Reagents / Q000737:chemistry; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000378:metabolism; D005557:Formaldehyde / Q000737:chemistry; D018628:Gene Dosage; D005786:Gene Expression Regulation; D007700:Kinetics; D011401:Promoter Regions, Genetic; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry / Q000378:metabolism; D035381:TATA-Binding Protein Associated Factors / Q000737:chemistry / Q000378:metabolism; D035181:TATA-Box Binding Protein / Q000737:chemistry / Q000378:metabolism*; D014157:Transcription Factors / Q000737:chemistry / Q000378:metabolism
chemical_list: D002843:Chromatin; D003432:Cross-Linking Reagents; D004268:DNA-Binding Proteins; C052130:GAL4 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D035381:TATA-Binding Protein Associated Factors; D035181:TATA-Box Binding Protein; D014157:Transcription Factors; D005557:Formaldehyde; D000251:Adenosine Triphosphatases; C073676:MOT1 protein, S cerevisiae